Submitted October 16, 2008
Accepted December 20, 2008
The structure of the ankyrin-binding site of 
-spectrin reveals how tandem spectrin-repeats generate unique ligand-binding properties
Paul R. Stabach, Ivana Simonovic, Miranda A. Ranieri, Michael S. Aboodi, Thomas A. Steitz, Miljan Simonovic, and Jon S. Morrow*
Department of Pathology, Yale University, New Haven, CT, United States
Department of Molecular Cellular Developmental Biology, Yale University, New Haven, CT, United States
Howard Hughes Medical Institute, New Haven, CT, United States
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, United States
* Corresponding author; email: jon.morrow{at}yale.edu.
Spectrin and ankyrin participate in membrane organization, stability, signal transduction, and protein targeting; their interaction is critical for erythrocyte stability. Repeats 14 and 15 of 
I-spectrin are crucial for ankyrin recognition, yet the way spectrin binds ankyrin while preserving its repeat structure is unknown. We have solved the crystal structure of the I-spectrin 14,15 di-repeat unit to 2.1 Å resolution, and find 14 residues critical for ankyrin binding that map to the end of the helix C of repeat 14, the linker region, and the B-C loop of repeat 15. The tilt (64o) across the 14,15 linker is greater than in any published di-repeat structure, suggesting that the relative positioning of the two repeats is important for ankyrin binding. We propose that a lack of structural constraints on linker and inter-helix loops allows proteins containing spectrin-like di-repeats to evolve diverse but specific ligand recognition sites without compromising the structure of the repeat unit. The linker regions between repeats are thus critical determinants of both spectrin's flexibility and poly-functionality. The putative coupling of flexibility and ligand binding suggests a mechanism by which spectrin might participate in mechanosensory regulation.
